1. Field of the Art
This invention relates to a DNA strand having an ability in biotechnological production of .alpha.-acetolactate decarboxylase (hereinafter called .alpha.-ALDCase) as produced by Enterobacter aerogenes IFO 13534 and to a yeast belonging to Saccharomyces cerevisiae transformed with the DNA strand so that its .alpha.-acetolactate (hereinafter called .alpha.-AL) producing ability is reduced.
2. Prior Art
Alcoholic beverages such as beer, sake, wine, etc., are generally produced by adding a yeast belonging to Saccharomyces cerevisiae to a starting material liquid for fermentation such as wort, fruit juice, etc., and subjecting the mixture to alcohol fermentation. In the fermentation process, the yeast will produce .alpha.-AL as the intermediate substance for biosynthesis of valine and leucine which are amino acids necessary for the growth of itself, and leak it inevitably out of the cell, namely into the fermented liquor. The .alpha.-AL which has thus become to exist in the fermented liquor will change spontaneously to diacetyl (hereinafter called DA) through the non-enzymatical reaction in the fermented liquor.
DA is a substance having strong objectionable odor called generally as "cooked odor" or "DA odor" and, in order to produce an alcoholic beverage excellent in flavor (namely without DA odor), the content of .alpha.-AL and DA in the fermented liquor is required to be decreased to a low level so that the total DA content will not finally exceed the discrimination threshold of DA odor in the alcoholic beverage (e.g. 0.05 to 0.1 mg/liter in the case of beer) even if .alpha.-AL may be all changed to DA.
While DA in the fermented liquor is converted to acetoin which is tasteless and odorless relatively rapidly in the co-presence of yeast, .alpha.-AL in the fermented liquor will not be changed by yeast, but it becomes decomposable with yeast only after it is changed to DA by non-enzymatical chemical reaction. However, since the reaction from .alpha.-AL to DA in the fermented liquor proceeds at a very slow rate, this reaction becomes the rate-limiting step, whereby the fermented liquor is required to be aged under the co-presence of yeast for a long time in order to obtain an alcoholic liquor with low content of .alpha.-AL and DA (namely without DA odor).
.alpha.-ALDCase is an enzyme having the property of converting .alpha.-AL to acetoin and has been known to be produced by various kinds of bacteria such as Enterobacter aerogenes, Bacillus licheniformis, Lactobacillus casei, Bacillus brevis, Enterobacter cloacae, Acetobacter bacteria (such as A. rancens, A. aceti, etc), etc. The principal enzymological properties of the .alpha.-ALDCase produced by Enterobacter aerogenes were investigated by the present inventors to obtain the results shown below.
Molecular weight: 28000.about.29000 PA1 Isoelectric point: pH 5.0.about.6.0 PA1 Optimal pH: 6.5.about.7.5 PA1 Optimal temperature: 40.degree..about.50.degree. C.
Thermal stability: stable to around 60.degree. C.
There is also a report about the enzymological properties of the .alpha.-ALDCase produced by Enterobacter aerogenes [European Journal of Biochemistry (Eur. J. Biochem.) 14 (1970) 133-137]